Epidermal cyclic GMP is increased in psoriasis lesions

Cyclic GMP levels in epidermis of normal subjects and of psoriatic patients were measured with a highly sensitive radioimmunoassay method. Technical improvements for the assay are 2-fold: (1) skin samples were frozen in vivo before biopsy and local injection of any anesthetic was avoided to overcome ischemia effect which could lower cyclic GMP artificially; (2) epidermis was microdissected to avoid contamination of dermis and keratin layers. The results show that on a per mg tissue dry weight basis the cyclic GMP levels are about 200 fmol in the involved lesional epidermis and 70 fmol in the uninvolved or normal epidermis. Similarly increases in the cyclic GMP levels in the lesional epidermis are observed when the data are expressed either on a DNA or protein basis. The cyclic GMP level in normal epidermis from nonpsoriatic subjects is the same as that in the uninvolved epidermis of psoriasis patients.     

Vasoactive intestinal polypeptide in allergic contact dermatitis: an immunohistochemical and radioimmunoassay study

Abstract Vasoactive intestinal polypeptide (VIP) is a neuropeptide with immunomodulatory properties. To elucidate the possible role of VIP in the pathophysiology of cutaneous contact hypersensitivity, we compared involved with uninvolved skin of allergic contact dermatitis (ACD) from nickel-allergic patients. Assays included quantification of VIP-immunoreactive (VIP-IR) nerve fibres and cells bearing immunoreactivity for VIP1 and VIP2 receptors in skin biopsy specimens, and of the concentration of VIP-like immunoreactivity (VIP-LI) in extracts of biopsy specimens. VIP-IR nerve fibres were found in the deeper part of the dermis close to sweat glands and hair follicles. No difference in the presence of VIP-IR nerve fibres was found between involved and uninvolved skin of ACD. VIP1 and VIP2 receptor immunoreactivity was seen on keratinocytes in the basal layer of the epidermis, with no difference between involved and uninvolved skin. Staining was also seen on vessel walls and mononuclear cells in the dermis. The highest staining intensity in the mononuclear cells was noted with the antibodies against the VIP2 receptor. While most of the mononuclear cells were stained in uninvolved skin, a minority of the cells showed a positive signal in involved skin. The concentration of VIP-LI in uninvolved skin was 1.53 ± 0.790 pmol/g and in involved skin 1.41 ± 0.735 pmol/g. It is concluded that there is no significant difference in either the distribution of VIP-IR fibres or the concentration of VIP-LI between involved and uninvolved skin of ACD. However, the number of dermal mononuclear cells showing VIP2 receptor immunoreactivity in skin of ACD was reduced. Received: 4 May 1998 / Received after revision: 12 October 1998 / Accepted: 16 October 1998     

Steroid hormone receptors in human melanoma

Summary Human melanomas were investigated for the presence of highaffinity estrogen-, gestagen-, and glucocorticoid-binding proteins. A statistically significant difference was found for mean estrogen receptor (ER) concentrations in melanomas of male versus female origin: female origin 37.6 (0–107) fmol/mg protein, male origin 3.9 (0–8.3) fmol/mg protein. No significant difference between sexes was found for gestragen receptors: 41.5 (0–194) fmol/mg protein for melanomas of female origin versus 99 (0–362) fmol/mg protein for male.Sucrose density gradient analyses revealed specific binding for both receptor types in the 4–5 S region as well as in the 8 S region. The binding affinities were in the same order of magnitude as reported for receptors found in typical steroid target organs.No significant difference in receptor values depending on sex was found for the glucocorticoid receptor: 19.2 (0–43) fmol/mg protein.     

Increased cholera toxin-, and forskolin-induced cyclic AMP accumulations in psoriatic involved versus uninvolved or normal human epidermis

Psoriatic involved epidermis reveals variously altered receptor-adenylate cyclase responses; among them the most prominent is defective beta-adrenergic adenylate cyclase response, which is normally the major receptor-adenylate cyclase system of human epidermis. It is known that activation of hormone-stimulated adenylate cyclase, a membrane-bound enzyme complex, requires functional coupling of at least 3 distinct subunits: 1) receptor subunit (R), 2) guanine nucleotide binding protein (G), and 3) catalytic subunit (C). The precise nature of the beta-adrenergic defect in the psoriatic epidermis, however, remains to be determined, especially in terms of G and C function. Using the involved and uninvolved skin from psoriatic patients, we investigated effects of cholera toxin (which monitors G-C interaction) and forskolin (which monitors C function) on the adenylate cyclase system of epidermis, which were compared with those of normal human epidermis. Both agents increased cyclic AMP levels of involved, uninvolved, and normal human epidermis. Marked accumulations were observed in the presence of cyclic nucleotide phosphodiesterase inhibitor, isobutyl-methylxanthine (IBMX); without the phosphodiesterase inhibitor, the effect of each agent was minimal. Comparison of the effects of cholera toxin revealed that the psoriatic involved epidermis accumulates much more cyclic AMP than the uninvolved epidermis (involved: 193 +/- 65; uninvolved: 117 +/- 54 pmoles/mg protein/5 h). Similarly forskolin-induced cyclic AMP accumulations of the involved epidermis were much more than those of uninvolved epidermis (involved: 374 +/- 152; uninvolved: 101 +/- 41 pmoles/mg protein/2 h). Those of normal human epidermis were not significantly different from those of uninvolved epidermis (cholera toxin: 99 +/- 36 pmoles/mg protein/5 h; forskolin: 84 +/- 22 pmoles/mg protein/2 h). Our results indicate that G and C function and their interaction is not defective (but rather increased) in the psoriatic involved epidermis. This suggests that the defective beta-adrenergic response of psoriatic involved epidermis reflects defective R or R-G interaction of the epidermal adenylate cyclase system.     

Pachydermoperiostosis: study of epidermal growth factor and steroid receptors

Pachydermoperiostosis is a rare osteo-cutaneous disease characterized by hypertrophy of bones and surrounding soft tissues. The cutaneous manifestations include coarsening of facial features, cutisverticis gyrata, digital clubbing, hyperhidrosis and scborrhoea. The pathogenetic mechanism of the disease is still debated, and proposed aetiological factors include genetic influences, anomalies in fibroblast activity, or alteration in peripheral blood flow. We studied a patient with the incomplete form of Pachydermoperiostosis. assessing epidermal growth factor receptor (EGF-R) and sex hormone steroid receptors (SR) in the affected skin, and also evaluating the urinary excretion of EGF. The results showed high levels of nuclear steroid receptors, increased cytosolic oestrogen receptors, and no detectable progesterone and androgen cytosolic receptors. EGF-R was also undetectable, and the urinary excretion of EGF was elevated. These findings suggest that the increased tissue sensitivity to circulating sex-steroids could induce enhanced tissue EGF/transforming growth factor α (TGF-α) production and utilization. The SR-EGF-R system could therefore be involved in determining hypertrophy of the affected tissues.     

Protein kinase C activity in normal and psoriatic cells: cultures of fibroblasts and lymphocytes

Protein kinase C (PKC) activity was measured in cultures of fibroblasts from biopsies of the involved and uninvolved skin of seven patients with psoriasis and from the skin biopsies of nine normal controls. PKC activity was significantly increased (P less than 0.005) in the particulate fraction of fibroblasts obtained from the involved areas of skin (450 +/- SEM 89 pmol/mg protein/3 min) and the uninvolved skin (394 +/- 94 pmol/mg protein/3 min) in psoriasis as compared to that of controls (103 +/- 24 pmol/mg protein/3 min). The soluble fraction of PKC activity was comparable in controls and in the fibroblasts obtained from involved areas and not significantly different from the values in fibroblasts from uninvolved skin. PKC activity was also measured in the soluble and particulate fractions of lymphocytes from 13 patients with psoriasis and from 14 normal controls. The PKC activity did not differ in the lymphocytes of patients with psoriasis from the controls in either the cytosolic or the membrane fractions. The increase in PKC activity as expressed at the membrane level of psoriatic fibroblasts may be related to an increase in sensitivity of these cells to hormones or growth factors involved in the regulation of their growth.     

A Case of Atrophoderma of Pasini and Pierini: Analysis of Glycosaminoglycan of the Lesional Skin

We report a case of atrophoderma of Pasini and Pierini. We determined the glycosaminoglycan content in the involved skin. Dermatan sulfate content in the involved skin (1.88 μg uronic acid/mg dry skin) was greater than that in the uninvolved skin (1.05 μg uronic acid/mg dry skin). No significant differences in hyaluronic acid, chondroitin sulfate or heparan sulfate content between involved and uninvolved skin were observed. These results suggest that abnormal metabolism of dermatan sulfate may be involved in the pathogenesis of atrophoderma; this pattern has been observed in systemic or localized scleroderma.     

cANBP与银屑病

本文采用竞争性蛋白质结合分析法测定了22例银屑病:患者受累与未受累皮肤的cAMP水平.同时应用了两种改进的皮肤切片技术:一是在样品中包括全部表皮和真皮,二是同时切取受累与未受累的皮肤样品,以避免由于缺血(Ischemic)作用而引起的人为差异.结果表明受累皮肤的CAMP水平明显低于未受累皮肤的CAMP水平.银屑病是与CAMP缺乏有关的.     

Psoriasis and vitamin A

Summary The vitamin-A status of 107 patient with psoriasis and 37 healthy controls was investigated. The mean serum level of retinol-binding protein (RBP) was normal in the 79 patients with chronic plaque psoriasis covering 25% or less of the skin surface. In the 28 patients with more extensive plaque lesions or pustular/ erythrodermic psoriasis, the mean serum RBP level was significantly lower than in the controls (P<0.05). The cutaneous concentrations of retinol (vitamin A₁), dehydroretinol (vitamin A₂) and carotenoids were measured in extracts of saponified shave-biopsy specimens of uninvolved and involved skin from 33 patients with plaque psoriasis. Their retinol values did not differ significantly from those found in control skin (mean, 252 ng/g), whereas the carotenoid levels in both uninvolved and involved skin were 25%–50% lower. In contrast, the dehydroretinol concentration was higher in the patients' involved skin (mean, 237 ng/g) than in their uninvolved skin (94 ng/g) and healthy control skin (70 ng/g; P<0.01). Although the origin of increased dehydroretinol levels in involved psoriatic skin is unknown, similar increments were observed in control epidermis in which proliferation had been induced by tape stripping. In 7 patients treated with oral etretinate (aromatic retinoid) for 2–3 weeks, the median retinol and dehydroretinol levels in involved skin increased by 107% and 212%, respectively; the vitamin-A concentrations in uninvolved skin did not change significantly. Oral treatment with -carotene/canthaxanthin raised the median carotenoid levels in uninvolved and involved skin by 170% and 610%, respectively, without significantly affecting the vitamin-A composition.     

LTA4 hydrolase in human skin: decreased activity,but normal concentration in lesional psoriatic skin

Leukotriene A₄ (LTA₄) hydrolase which transforms LTA₄ into the proinflammatory compound LTB₄ has been identified in human epidermis. The purpose of this study was to investigate the potential role of this enzyme in psoriasis, in which LTB₄ is present in biologically active concentrations. The concentration and activity of LTA₄ hydrolase was determined in normal skin and in matched samples of involved and uninvolved psoriatic skin. The enzyme content was determined using an affinity-purified antibody. This antibody was also used for immunohistochemical staining of skin biopsies. Immunohistochemically LTA₄ hydrolase was localized predominantly in the basal and spinous layers in normal skin and in involved and uninvolved psoriatic skin. The LTA₄ hydrolase content varied between 2.8 and 3.1 μg enzyme/mg protein and was found to be similar in normal and psoriatic skin, involved as well as uninvolved. In contrast, the activity of the enzyme was decreased significantly in involved psoriatic skin (9.9±2.1 μg LTB₄/mg enzyme per min) compared with matched uninvolved psoriatic skin (16.4±3.5 μg LTB₄/mg enzyme per min), but was decreased only insignificantly compared with normal skin (12.4±1.8 μg LTB₄/mg enzyme per min). It was found that the conversion of LTA₄ to LTB₄ results in inactivation of LTA₄ hydrolase activity. This finding is compatible with the idea that the decreased LTA₄ hydrolase activity in involved psoriatic skin reflects transcellular LTB₄ formation in vivo. In peripheral lymphocytes the enzyme content was 1.3±0.3 μg enzyme/mg protein in normal lymphocytes and 1.4±0.3 μg enzyme/mg protein in psoriatic lymphocytes, which was significantly lower than in the skin. In contrast, the specific LTA₄ hydrolase activities in normal and psoriatic lymphocytes (23.4±1.3 and 21.3±1.7 μg LTB₄/mg enzyme per min) were significantly higher than in normal skin. These findings may indicate the existence of LTA₄ hydrolase isoforms in human lymphocytes and human skin.     

Gene expression analysis of melanocortin system in vitiligo

BACKGROUND: The melanocortin system in the skin coordinates pigmentation and immune response and could be implicated in the pathogenesis of vitiligo. OBJECTIVES: We aimed to analyze changes in expression of genes involved in skin pigmentation (melanocortin system and enzymes involved in melanin synthesis). METHODS: With quantitative RT-PCR we measured the mRNA expression levels of eight genes from the melanocortin system and two enzymes involved in melanogenesis. RNA was extracted from both lesional and non-lesional skin of vitiligo patients and in non-sun-exposed skin of healthy subjects. RESULTS: POMC (proopiomelanocortin) expression was lower in lesional skin compared to non-lesional skin. Expression of melanocortin receptors was increased in unaffected skin of vitiligo patients compared to healthy subjects and decreased in lesional skin compared to uninvolved skin of vitiligo patients, the differences were statistically significant in the cases of MC1R (melanocortin receptor 1) and MC4R (melanocortin receptor 4). TRP1 and DCT genes were down-regulated in lesional skin compared to non-lesional vitiligo skin or skin of healthy controls and up-regulated in uninvolved vitiligo skin compared to healthy control samples. In non-lesional skin, POMC expression was not elevated, possibly indicating that systemic influences are involved in up-regulation of MC receptor genes. Decreased expression of the analyzed genes in the lesional skin is not surprising, but statistically significant increased expression of studied genes in non-lesional skin from vitiligo patients is not described previously. CONCLUSION: In our mind, up-regulation of melanocortin system in non-lesional skin could be systemic compensation to restore normal pigmentation in lesions.     

Altered [125I]epidermal growth factor binding and receptor distribution in psoriasis

Stimulation of growth and differentiation of human epidermis by epidermal growth factor (EGF) is mediated by its binding to specific receptors. Whether EGF receptors primarily mediate cell division or differentiation in hyperproliferative disease such as psoriasis vulgaris is unclear. To study the pathogenesis of psoriasis, 4-mm2 punch biopsy specimens of normal, uninvolved, and involved psoriatic skin were assayed for EGF receptors by autoradiographic, immunohistochemical, and biochemical methods. Using autoradiographic and immunohistochemical methods, basal keratinocytes were found to contain the greatest number of EGF binding sites and immunoreactive receptors as compared to the upper layers of the epidermis in both normal epidermis and psoriatic skin. No EGF receptor differences between normal and psoriatic epidermis were observed in this layer. In the upper layers of the epidermis, a 2-fold increase in EGF binding capacity was observed in psoriatic skin as compared with normal thin or thick skin. Biochemical methods indicated that [125I]EGF binding was increased in psoriatic epidermis as compared with similar thickness normal epidermis when measured on a protein basis. Epidermal growth factor was shown to increase phosphorylation of the EGF receptor in skin. EGF receptors retained in the nonmitotic stratum spinosum and parakeratotic stratum corneum may reflect the incomplete, abnormal differentiation that occurs in active psoriatic lesions. Alternatively, retained EGF receptors may play a direct role in inhibiting cellular differentiation in the suprabasal layers.     

Intranuclear androgen and cytosolic receptor concentrations in the axillary skin of osmidrosis

Summary 5-Dihydrotestosterone (DHT) and testosterone were measured by radioimmunoassay in the crude nuclear and cytoplasmic fractions of the axillary skin of both male and female patients with osmidrosis and the levels compared with those of nongenital skin. The intranuclear levels of DHT were 1.44±0.22 and 1.77±0.38 pg/g DNA in men and women, respectively. Those of testosterone were about 10% of DHT levels. In the skin of nontarget regions nuclear DHT was much scarcer or undetectable. Cytosolic androgen receptors in isolated apocrine glands were also measured using ³H-R1881 as a ligand. Typical androgen receptors were present in all of eight patients (K_D=1.32 ±0.24×10^–9M, B_max=10.3±0.51 fmol/mg protein). Neither the intranuclear androgen concentrations nor the cytosolic androgen receptor levels were significantly different between the two sexes. These data indicate clearly that the apocrine gland of patients with osmidrosis is a typical androgen target organ, irrespective of sex, and suggest that nuclear DHT in the axillary skin of women is derived from not only testosterone but also other precursors, especially in consideration of the very low serum concentrations of testosterone in females.     

Characterization of the glucocorticoid receptor in human epidermis and dermis

The binding of [³H]triamcinolone acetonide to cytosol proteins in human epidermis and dermis has been characterized. Qualitatively the binding in both cytosols was very similar. The binding was of high affinity (dissociation constant = 1·07 × 10^?9 mol/l and 1·36×10^?9mol/l for epidermal and dermal binding proteins respectively) and low capacity (27·3 fmol/mg protein and 53·0 fmol/mg protein for epidermal and dermal binding proteins respectively). The binding protein was labile m heating at 40°C and proteolytic enzymes. Glucocorticoids showed an affinity which approximated to their therapeutic potency in both epidermis and dermis and non-glucocorticoids had little or no binding affinity. Quantitatively more than 90% of the binding protein in whole skin was present in the dermis and less than 10% in the epidermis, which approximates to a difference in tissue mass. There were 12·O× 10¹⁰ receptors in a disc of whole skin of 1 cm² surface area of which 11·0×10¹⁰ were in the dermis, 0·95 × 10¹⁰ in the epidermis with 1706 and 1214 receptor sites for epidermal and dermal cells respectively. Receptor concentration is highest in dermis when expressed per cytosol protein and in epidermis when expressed per DNA. It is concluded that the binding protein characterized in epidermis and dermis is the physiological glucocorticoid cytosol receptor. In the absence of qualitative differences between the epidermal and dermal receptors quantitative differences may help in the development of less toxic therapeutic agents.     

Epidermolysis bullosa acquisita

Four patients with the clinical picture of epidermolysis bullosa acquisita were investigated. Biopsies were taken from the involved and uninvolved areas of the skin and the immunohistochemical and microscopic changes were studied. Direct immunofluorescence showed deposition of IgG and C3/4 in a linear or notched pattern along the epidermal basement membrane in both the involved and the uninvolved skin. In addition IgA (3/4), IgM (1/4), C4 (3/4) and properdin (3/4) could be detected. Indirect immunofluorescence revealed the presence of circulating antibodies against inter alia the epithelial basement membrane zone in one patient. Routine electron microscopy showed that the blister was situated in the dermis leaving the basal lamina in the roof of the blister. With immunoelectron microscopy using peroxidase-labelled antibody the in vivo deposition of IgG was observed just beneath the basal lamina in the dermis of both the perilesional and the uninvolved skin. These observations show that epidermolysis bullosa acquisita is a distinct entity, in which autoimmune mechanisms might possibly play a role.     

Decreased protein kinase C activity in psoriatic versus normal epidermis

To study the possible involvement of protein kinase C in psoriasis, we determined the activity of this enzyme in involved and uninvolved epidermis from psoriasis patients as well as in normal epidermis from controls. Protein kinase C activity was measured in the 100,000 g supernatants and in the detergent-solubilized particulate fractions of tissue homogenates after partial purification by DEAE-cellulose chromatography. The total enzyme activities per gram protein for normal, involved, and uninvolved epidermis were 127 +/- 11.9 mU/g, 64.7 +/- 8.6 mU/g, and 88.5 +/- 14.4 mU/g, respectively. The respective values for total enzyme activity per mg DNA were 7.04 +/- 0.48 mU/mg, 4.28 +/- 0.54 mU/mg, and 5.02 +/- 0.66 mU/mg. By both data bases, protein kinase C activity was statistically lower in the psoriatic skin biopsies than in those from control persons, whereas no significant difference was found between involved and uninvolved epidermis from psoriasis patients. We hypothesize that alterations in protein kinase C-mediated processes due to decreased protein kinase C activity may play a significant role in the pathophysiology of psoriasis.     

Downregulation and altered spatial pattern of caveolin-1 in chronic plaque psoriasis

BACKGROUND: Caveolin-1 is a key structural and functional protein for plasmalemmal invaginations termed caveolae. Caveolin-1 is known to modulate multiple signal-transducing pathways involved in cell differentiation and proliferation. Psoriasis is viewed as a multifactorial pathology characterized by keratinocyte hyperproliferation and abnormal cell maturation. We hypothesized that loss of caveolin-1 within epidermal keratinocytes may contribute to the development and/or progression of the psoriatic phenotype. OBJECTIVES: To examine the expression and spatial distribution of caveolin-1 in skin biopsies from normal subjects and in patients with psoriasis. METHODS: Using immunohistochemical methods caveolin-1 protein expression was assayed in two independent patient groups. Firstly, a retrospective analysis was conducted on archival skin samples obtained from nine normal subjects and from involved tissue of 12 patients with psoriasis. Following this, a prospectively designed study was conducted in 10 further patients with active psoriasis and involving caveolin-1 staining of biopsy tissue from the uninvolved, advancing edge and lesional skin tissue from within the same subject. RESULTS: In normal skin or uninvolved skin from psoriasis patients intense caveolin-1 staining was present throughout full-thickness epidermis. In 20 of the 22 patient cases (combined retrospective and prospective samples) caveolin-1 protein was significantly reduced and consistently showed very weak or absent staining within the hyperproliferative basal cell layers of the psoriatic plaque (P < 0.002 for retrospective archival study and P < 0.01 for prospectively designed study). Comparisons between caveolin-1 staining in uninvolved tissue and at the advancing edge of a migrating plaque were more equivocal (P > 0.05). CONCLUSIONS: The findings of this study are consistent with a downregulation of caveolin-1 that may serve as an aetiological factor in the development and/or progression of psoriasis. Further mechanistic investigations are required with the potential that caveolin-1 protein may be a novel target for therapy of psoriasis.